Tests are listed in alphabetical order

Acid Detergent Fiber (ADF)

Fiber (Acid Detergent) and Lignin in Animal Feed: AOAC Official Method 973.18 (1996)

Modification includes use of sea sand for filter aid.

Acid Detergent Insoluble Protein (AD-ICP)

Fiber (Acid Detergent) and Lignin in Animal Feed: AOAC Official Method 973.18 (1996)

Protein (Crude) in Animal Feed: Combustion Method. AOAC Official Method 990.03 (2012)

Amino Acids

High performance liquid chromatography (HPLC) separates amino acids using ion exchange chromatography. Following separation, the amino acids are derivatized with trione, enabling their detecting at 560 nm.

Gehrke, C. W., L. Wall Sr., J. Absheer, F. Kaiser, and R. Zumwalt. 1985. Sample preparation for chromatography of amino acids: Acid hydrolysis of proteins. J. Assoc. Off. Anal. Chem. 68:811–821. https:/ / .1093/ jaoac/68 .5 .811.

Ammonia CPE

Ammonium-N determination by distillation, Recommended Methods of Manure Analysis. ch4 sec1: 25 (2003)

Modification using Kjelsorb reagent.


Ash of Animal Feed: AOAC Official Method 942.05 (1996).

Sample is extracted in water, filtered and measured by way of Corning 926 Chloride Analyzer.
Corn Silage Processing Score (CSPS)

Particle Size, Fragmentation Index, and Effective Fiber: Tools for Evaluating the Physical Attributes of Corn Silages. Proc. Four State Dairy Nutrition pg 211-220 2005 Mertens, David R., 2005

Crude Fiber

Fiber (Crude) in Animal Feed and Pet Food (Fritted glass crucible method): AOAC Official Method 978.10 (1979)

Modification includes use of sea sand for filter aid.

Crude Protein
Protein (Crude) in Animal Feed: Combustion Method. AOAC Official Method 990.03 (2012)
Dry Matter - Dairy Liquid

Solids (Total) in Milk. AOAC Official Method 990.20

Dry Matter - Forages

Partial dry matter is determined by drying < 55°C in a forced-air oven to < 6% residual moisture, followed by drying for 3 hrs at 105°C according to NFTA Method 2.1.4 to determine residual dry matter.

E.Coli and Coliform

AOAC Official Method 2014.05 Enumeration of coliform/E.coli in feed

Fat (Ether Extract)

Diethyl Ether Extract

Default Method
Fat (Crude) or Ether Extract in Animal Feeds, AOAC Official Method 920.39

Automated with Foss Soxtec 2047 instrument with the use of diethyl ether

Petroleum Ether Extract

Official Method of the Distillers Grain Technology Council
Fat (Crude) or Ether Extract in Animal Feeds,  AOAC Official Method 920.39

Automated with Foss Soxtec 2047 instrument with the use of petroleum ether

Fat (Acid Hydrolysis)
(As requested or with non-dairy liquid samples)
Determination of total fat in feed and feed ingredients using the SoxCap 2047 in combination with Soxtec extraction systems, Foss Analytical AB Soxtec System Application Note AN3902 (2006).
Fatty Acid (TFA)
Fatty Acid Analysis in Animal Feeds: Method workshop from 30th Annual MW AOACI Meeting and Exposition. Kraft, J. and Palmquist, D. (Modification using a packed column instead of capillary column).
Forage Particle Size
Method of Determining and Expressing Particle Size of Chopped Forage Materials by Screening, ASAE Forage Harvesting and Utilization Committee, ANSI/ASAE S424 June 24, 1988.
Grain Particle Size
Measuring and defining particle size of feedstuffs. In: First International Symposium on Particle Size Reduction in the Feed Industry. Kansas State University, Manhattan, KS. Behnke, K. 1985.
In Vitro Starch Digestibility - 7 Hour

Richards, C.J., J.F. Peterson, R.A. Britton, R.A. Stock, and C.R. Krehbiel. 1995. In vitro starch disappearance procedure modifications. Anim. Feed Sci. and Tech. 55:35-45.
Modifications include using Goering and Van Soest (1970) method with inoculum made from a composite of strained ruminal fluid and strained fluid from blended ruminal solids with an equivalent volume of reduced media


A Method for Partitioning Neutral Detergent-Soluble lactose, J. Sci. Food Agric. (1999),#79: 2079 


Nutritional Importance and Classification of Food lactose, Plant Polymeric lactose. (1993), Asp N-G, Royal Society of Chemistry, Cambridge UK,  pp 121-126 


Colorimetric Method for Determination of lactose and Related Substances, Anal. Chem. (1956), 28:350 


Neutral Detergent-Soluble lactose Nutritional Relevance and Analysis,   Hall, M.B., Lab Manual(University of Florida) Bulletin 339 


Method for determination water soluble lactose Derias, R.E. 1961. J. Sci. Fd. Agric. 12:152  


Fiber (Acid Detergent) and Lignin in Animal Feed: AOAC Official Method
973.18 (1996) (Modification includes use of Sea Sand for filter aid as needed, use of Whatman GF/C filter paper to collect residue, and holding crucibles in beakers to cover fiber with 72% sulfuric acid for full time required).

Metals in Plants

AOAC Official Method 953.01 16th edition, Ch3 pg 2.

Dry Ash

Phosphorus, Potassium, Calcium, and Magnesium (Short Mineral Analysis) Wisconsin Soil Testing and Plant Analysis Procedures No. 6 Soil Fertility Series 1970 Revised 1977.

Microwave Digestion

50% Nitric Acid in High Energy CEM MarS Express microwave diluted to 10% then analyzed on ICP.

Mineral Mix

.5 grams of sample, 10 mls of HCl to dryness then 10 mLs of Aquaregia for 10 minutes on a hot plate, diluted to 100mL of water then analyzed by ICP.

Determination of Sulfur in feed materials using ICP-OES and microwave digestion; Mullins S., Bartos J., Newlon N., Office of Indiana State Chemist. May 2007.

Mojonnier Fat
Mojonnier method, Standard Methods for the Examination of Dairy Products; 16th edition ch15F pp474-479 (1992).
Mold/Yeast Count and Identification

Aerobic Plate Count

Standard Methods for the Examination of Water and Wastewater, 19th (1995),#9215 B

Heterotrophic Plate Count

Standard Methods for the Examination of Water and Wastewater, 19th (1995),#9215 A

Potato Dextrose Mold Analysis

Molds and Mycotoxins in Foods (Shortcourse).,University of Nebraska-Lincoln, March 17-19 2009. Dept of Food Science and Technology

Petrifilm for Yeast and Mold Count Analysis

AOAC Official Method.,2014.05”Enumeration of Yeast and Mold in Food



Multitoxin Mycotoxin Detection Method by LC-MS/MS
S. M. Sulyok, F. Berthiller, R. Krska, R. Schuhmacher. 2006. Development and validation of a liquid chromatography/tandem mass spectrometric method for the determination of 39 mycotoxins in wheat and maize. Rapid Commun. Mass Spectrom. 20: 2649-2659.

Zhang, K., M.R. Schaab, G. Southwood, E.R. Tor, L.S. Aston, W. Song, B. Eitzer, S. Majumdar, T. Lapainis, H. Mai, and K. Tran. 2017. A collaborative study: Determination of mycotoxins in corn, peanut butter, and wheat flour using stable isotope dilution assay (SIDA) and liquid chromatography–tandem mass spectrometry (LC-MS/MS). J. Agric. Food Chem. 65:7138-7152.

Neutral Detergent Fiber (aNDF)
Amylase-Treated Neutral Detergent Fiber in Feeds AOAC Official Method 2002.04 2005(Modification includes use of Sea Sand for filter aid and Whatman GF/C filter paper for residue collection).
Neutral Detergent Fiber Digestibility (NDFD)

Fermented in flasks using the method of Goering and Van Soest (1970) with the modification that a composite inoculum containing strained ruminal fluid and blended ruminal solids. Residue analyzed by NDF procedure.

Neutral-Detergent Insoluble Protein (ND-ICP)
Residue from the NDF method (minus sodium sulfite) is collected by use of Whatman 541 (nitrogen-free) paper or Whatman GF/C filter paper before analysis for nitrogen by Crude Protein method.

Screening Analysis

Blue-Violet for Nitrate Ion. Journal of Chem. Educ Grotz, Leonard C.; Vol. 50, No. 1 Jan. 1973, p 63(Diphenylamine used on dried ground sample to screen for nitrate presence).

Quantitative Analysis

Nitrate in Forages.  AOAC Official Method 986.31 16th Edition, Chp14 pg 5. (Modified to include boric acid, silver sulfate, and sulfamic acid in the extracting solution).

Non-Protein Nitrogen (NPN) and Urea

Urea and Ammoniacal Nitrogen in Animal Feed, AOAC Official Methods of Analysis (2002)941.04 4.3.01


Ammonium-N determination by distillation, Recommended Methods of Manure Analysis. (2003),ch4 sec1: 25


Nitrogen(Ammonia) in Water,.AOAC Official Methods of Analysis (1995),973.49, ch11 pp9-10


EPA 350.2

AOCS Official Method., BA 9-58. 

Physically Effective Neutral-Detergent Fiber (peNDF)

D. R. Mertens (1986, 1997) “proposed that physically effective NDF could be determined using particle size separation. Based on the size of particles passing the rumen and those found in the feces, he assumed that any fiber less than 1.18mm would not stimulate chewing activity. Thus, it may be possible to measure peNDF directly by determining the aNDF in particles <1.18mm and subtracting this from the total aNDF to obtain peNDF.”

The CSPS also takes into account how well the corn kernels have been processed by using total starch analysis in the separated material in order to predict the digestibility of the corn silage. Starch is a mixture of amylose and amylopectin. These are both complex carbohydrate polymers of glucose making starch a glucose polymer as well, regardless of the ratio of amylose:amylopectin.(Wikipedia) With the use of a buffer solution and proper enzymes, the starch can be degraded into glucose components. When accounting for free glucose, the total glucose can be used to determine the total starch of the samples.

A neutral detergent solution is used to dissolve the easily digested pectins and plant cell contents (protected sugars and lipids), leaving a fibrous residue (aNDF) that is primarily cell wall components of plants (cellulose, hemicellulose, and lignin). Detergent is used to solubilize the proteins and sodium sulfite also helps remove some nitrogenous matter. EDTA is used to chelate calcium and remove pectins at boiling temperatures. Triethylene glycol helps to remove some nonfibrous matter from concentrate feeds, and heat stable alpha-amylase is used to remove starch. Two additions of amylase (one during refluxing and one during filtration) have been observed to aid NDF analysis and minimize filtering difficulties. Heat stable amylase is used in hot solutions to inactivate potential contaminating enzymes that might degrade fibrous constituents.





Forage Fiber Analysis (Apparatus, reagents, procedures, and some applications): Goering, H.K.; Van Soest, P.J. (Modified per Mertens, D.R.) USDA Agricultural Research Service. Handbook Number 379(1970)(Mertens 1992)


Methods for Dietary Fiber, Neutral Detergent Fiber Non-Starch Polysaccharides in Relation to Animal Nutrition: Van Soest, P.J, Robertson, J.B, Lewis, B.A, J. Dairy Science 74:3583-3597 (1991)


Critical Conditions in Determining Detergent Fiber: Mertens, D.R. Proceedings of NFTA Forage Analysis Workshop pC1-C8 (1992)


Amylase-treated Neutral Detergent Fiber in Feeds: AOAC 2002.04


Determination of Starch in Large Particles Ro-Tap Method: Mertens, D.R. US Dairy Forage Research Center (2001)


Dietary Starch in Animal Feeds and Pet Food: AOAC 2014.10

Bach Knudsen—Journal of Animal Feed Science Technology 67(1997) pp319-338

Analytical Methods Guide, Orion Research, May 1977.
Determination of Prolamin Proteins in Corn. UW Grain Evaluation System
Workshop 6/2/09
Protein Dispersibility Index (PDI)
AOCS Official Method., BA 10-65.
Protein Digestibility

Development of an In Vitro Intestinal Digestibility Assay for Ruminant Feeds – Proceedings for 75th Cornell Nutrition Conference for Feed Manufacturers pg 190-202  Ross, D.A. 2013

Protein Solubility

Evaluation of protein solubility as an indicator of overprocessing of soybean meal. Poult. Sci. 69:76-83 1990 Araba, M. and N.M. Dale

The potassium hydroxide protein solubility test. NRN, LLC June 28, 2019

Direct communication with Nelson Ruiz. 2019 and 2020

Ross RUP (16 hours)
Development of an In Vitro Intestinal Digestibility Assay for Ruminant Feeds – Proceedings for 75th Cornell Nutrition Conference for Feed Manufacturers pg 190-202 Ross, D.A. 2013

Acid Hydrolysis Starch

Determining Corn Germ and Pericarp Residual Starch by Acid Hydrolysis Cereal Chemistry 86(2):133-135 Vidal, B.C Jr., Rausch, K.D., Tumbleson, M.E., Singh, V.

Dietary Starch

Starch Analysis in Animal Feed: Method workshop from 30th Annual MW AOACI Meeting and Exposition. Method that is undergoing AOAC collaborative study. (Modification: Glucose analysis completed on YSI 2700 Select Biochemistry analyzer instead of using GOPOD).

Sugar - Ethanol Soluble Carbohydrates (ESC)

Feeds are extracted in 80% ethanol and ESC are determined using sucrose as the standard carbohydrate

Hall, M. B., W. H. Hoover, J. P. Jennings, and T. K. Miller Webster. 1999. A method for partitioning neutral detergent-soluble carbohydrates. J. Sci. Food Agric. 79:2079–2086.

Sugar - Water Soluble Carbohydrates (WSC)
Extraction procedure adapted from Derias, R.E. 1961. J. Sci. Fd. Agric. 12:152 (Method for determination water soluble carbohydrates).  Detection procedure from Neutral Detergent-Soluble Carbohydrates Nutritional Relevance and Analysis, Hall, M.B., Lab Manual(University of Florida) Bulletin 339
Volatile Fatty Acids (VFA)
“Determination of Volatile Carboxylic Acids (C1-C5i) and Lactic Acid in Aqueous Acid Extracts of Silage by High Performance Liquid Chromatography”  Journal of Sci. Food Agric 35pp1178-1182 (1984) Modification by Dr. Richard Muck, U.S. Dairy Forage Research Center, Madison, WI.

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